ON THE DISTRIBUTION OF LIGANDS WITHIN THE ASYMMETRIC CHAPERONIN COMPLEX, GROEL(14)CENTER-DOT-ADP(7)CENTER-DOT-GROES(7)

In the presence of MgATP or MgADP the E. coli chaperonin proteins, GroEL and GroES, form a stable asymmetric complex with a stoichiometry of two GroEL(7):one GroES(7): seven MgADP. The distribution of the ligands between the two heptameric GroEL rings is crucial to our understanding of the mechanism of chaperonin-assisted folding, being either cis (i.e. [GroEL(7) . MgADP(7) . GroES(7)]-[GroEL(7)]) or trans (i.e. [GroEL(7) . MgADP(7)]-[GroEL(7) . GroES(7)]. On the basis of cross-linking experiments with andazido-ATP and the heterobifunctional reagent, N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP), it was suggested that GroES and MgADP are bound to the same GroEL ring which resists proteinase K digestion [Nature 366 (1993) 228-233]. However, we find that the SPDP-promoted cross linking of GroES and GroEL occurs in the absence of Mg2+, ADP or ATP, which are required for the formation of the asymmetric complex, Cross-linking is shown to occur only when the SPDP-modified GroES is co-precipitated with GroEL by trichloracetic acid, Furthermore, there are structural grounds for questioning whether SPDP can crosslink, in a physiologically relevant manner, an amino group of GroES with any of the cysteinyl groups of GroEL.